1-r-2, 4-dinitropyrroles



United States Patent 3,256,295 1-R-2,4-DINITROPYRROLES George Karmas,Bound Brook, N.J., assiguor to Ortho Pharmaceutical Corporation, acorporation of New Jersey No Drawing. Original application Nov. 5, 1964,Ser. No. 409,277. Divided and this application Sept. 7, 1965,

Ser. No. 485,544

1 Claim. (Cl. 260309) l IL in which R is a substituent selected from thegroup consisting of alkyl groups selected from the group consisting ofn-propyl, n-butyl, iso-butyl, sec-butyl, n-amyl, isopentyl, sec-pentyland n-hexyl, haloalkyl groups selected from the group consisting of'ybromopropyl and B-bromobutyl, acetonyl, e -dihydroxypropyl, fl,'-diacetoxypropyl, l-(N-morpholino carbonyl methyl), piperazino groupsselected from the group consisting of 1-(N methyl-N -piperazino carbonylmethyl) and 1-(N -acetyl- N -piperazino carbonyl methyl), and1-(2-methyl-5-nitroimidazolyll-ethyl) Examples of 2,4-dinitropyrrolescoming within the scope of the above formula are: l-(y-bromopropyl)-2,4-dinitropyrrole, 1-acetonyl-2,4-dinitropyrrole, l-isopentyl-2,4-dinitropyrrole, 1-(/3,-y-dihydroxypropyl) 2,4 dinitropyrrole andl-(N-morpholino carbonyl methyl)-2,4-dinitropyrrole.

The compounds of the present invention have antimicrobial activity andmore specifically are effective against Trichomonair foetus, a parasiticprotozoan that infects the uterus of animals and causes abortion incattle. The high order of trichomonodicidal activity possessed by thecompounds of the present invention is quite surprising as the closelyrelated 1-alkyl-2,S-dinitropyrroles and 1-alkyl-3,4dinitropyrroles haveno useful activity.

It is an object of the present invention to provide new compounds havingtherapeutic utility in the treatment of animals infected withTrichomonas foetus.

It is also an object of this invention to provide a new method for thepreparation of 1-substituted 2,4-dinitropyrroles.

Heretofore it has been proposed to prepare N-alkylpyrroles by reactingpyrrolewith potassium metal in an organic solvent such as a low boilinghydrocarbon and reacting the resulting N-potassiumpyrrole with an alkylhalide for example, methyl iodide. The latter reaction is carried outeither by heating the ingredients together in a sealed tube at 120-130C., or by refluxing in a low Patented June 14, 1966 boilingorganicsolvent. By this method, N-methylpyrrole can be obtained inyields around 25-50%. A considerable amount of pyrrole is converted toa-methylpyrrole, which side reaction is primarily responsible for therelatively low yield of the desired N-alkyl product.

It has also been proposed to react an alkali metal pyrrole in anhydrousliquid ammonia with an organic halide. Under such conditions, highyields have been obtained, but the anhydrous liquid ammonia solvent ismore difficult to handle than a solvent having a lower vapor pressure.Moreover, the alkylation reaction, at liquid ammonia temperature,requires several hours for completion.

In accordance with the present invention, the alkylation of2,4-dinitropyrroles is effected in dimethylformamide which is a superiorsolvent for this reaction. In general, the alkali metal salt of2,4-dinitropyrro1e and an excess of the ankylating agent, which may be achloride, bromide, iodide sulfate, sulfonate, etc., are heated indimethylformamide for varying periods of time (determined by thereactivity of the alkylating agent). The reaction product is isolatedand purified by conventional procedures of extraction, distillation,recrystallization, etc. In those instances where the grouping introducedat the l-position of the pyrrole contains a functional group, furtherreactions may be performed on this group. For example, an ester may behydrolyzed to a carboxylic acid and the latter may further be convertedto an acid chloride, from which a wide variety of derivatives may beprepared.

The following examples will serve to illustrate more fully the method ofpreparing the novel compounds of the present invention.

Example I.1-butyl-2,4-dinitr0pyrr0le A mixture of 5 grams (0.0275 mole)of the sodium salt of 2,4-dinitropyrrole, 8 milliliters of dibutylsulfate, and 20 milliliters of dimethylformamide is heated to a gentleboil and after one minute is cooled and poured with stirring into amixture of 200 milliliters of water, grams ice, and 15 grams of sodiumcanbonate layered with 150 milliliters of ether. This mixture is shakenvigorously and then the layers are separated. The ether solution isWashed with 100 milliliters of 5% aqueous sodium carbonate, dried withanhydrous magnesium sulfate, concentrated and then distilled. Thel-butyl2,4-dinitropyrrole is a pale yellow oil which distills at -135 C.at 0.2 mm.

Unalkylated 2,4-dinitropyrrole may be recovered from the aqueous sodiumcarbonate solutions by acidification and extraction with ether.

Dimethyl sulfate, diethyl sulfate, and dipropyl sulfate have also beenreacted with the sodium salt of 2,4-dinitropyrrole following thisprocedure.

Example II.1-isommyl-2,4-dinitr0pyrr0le A mixture of 5 grams (0.0275mole) of the sodium salt of 2,4-dinitropyrrole, 6 milliliters of isoamylbromide,

' and 10 milliliters of dimethylformamide is heated under reflux for twohours. The reaction mixture is worked up as described for the l-butylanalog (Example I), and

- 3 distillation afford 1-isoamyl-2,4-dinitropyrrole as a pale yellowoil which distills at 135140 C. at 0.1 mm.

Example III.1-propyl-2,4-dinitr0pyrr0le A mixture of 38.6 grams (0.215mole) of the sodium salt of 2,4-dinitropyrrole, 3O milliliters of propylbromide and 30 milliliters of dimethylformamide is heated in a sealedtube at 135-140" C. for 3 /2 hours. After cooling, the tube is openedand the reaction mixture is Worked up as described in Example I above.Distillation affords 24.7 grams of 1-propyl-2,4-dinitropyrrole. Thissolid product is recrystallized from an ether-ligroin mixture to give20.2 grams (47.2% yield) of pale yellow prisms, melting point 5657 C.

From the aqueous sodium carbonate solutions, a 35% recovery of2,4-dinitropyrrole is elfected an acidification with hydrochloric acidand extraction with ether.

Example IV.-1-(2,3-dihydr0xypropyl) 2,4 -d ini tropyrrole A mixture of 5grams (0.0275 mole) of the sodium salt of 2,4-dinitropyrrole, 6milliliters of a-glyceryl monochlorohydrin, and 20 milliliters ofdimethylformamide is heated under reflux for fifty minutes, cooledslightly, and then concentrated under vacuum to remove the solvent. Theviscous concentration residue is leached with three BOO-milliliterportions of boiling ether, decanting each time from the insolubleresidue. The combined ether solution is concentrated and the oilyresidue is distilled to afford the crude product as a viscous oil whichboils at 200-210 C., at 0.01-0.02 mm. It solidifies on standing and isrecrystallized from ethyl acetate to give 2.4 grams (37.8% yield) of1-(2,3-dihydroxypropyl)-2,4-dinitropyrrole, small cream prisms ofmelting point 111-112 C.

This procedure is generally suitable for the preparation of watersoluble 1-substituted-2,4-dinitropyrroles because it avoids the loss ofthese in aqueous wash solutions. It has been applied to the synthesis ofthe corresponding l-(B-hydroxyethyl) and l-(y-hydroxypropyl) analogs.

Example V.1-(my-diacetoxypropfl) 2,4-dinitrpyrr0le A mixture of 7.3grams of 1-(B,'y-dihydroxypropyl)- 2,4-dinitropyrrole (Example IV) and60 milliliters of acetic anhydride is heated under reflux for ninetyminutes, cooled slightly, and then concentrated under vacuum to removethe excess of acetic anhydride. The syrupy residue is dissolved in 20milliliters of boiling ethyl acetate and the solution is stored at 0 tocrystallize 8.4 grams of pale yellow granules which melt at 109l10. Asecond crop of 1.0 gram is isolated by concentration of the motherliquor. The total of 9.4 grams represents a yield of 94%.

Example Vl.1-carb0xymethyl-2,4-dinitr0pyrr0le To a cold C.) solution of5.2 grams (0.0214 mole) of 1-carbethoxymethyl-2,4-dinitropyrrole(prepared according to the process of Example II, from ethylchloroacetate) in 150 milliliters of methanol is added 0.855 gram(0.0214 mole) of sodium hydroxide in 10 milliliters of water. To theresulting paste is added 200 milliliters of water and this mixture iswarmed at 45 C. for 10 minutes. After it has been cooled to 0 C., thissolution is acidified with hydrochloric acid and the precipitatedcarboxylic acid is filtered off, washed on the filter with cold water,and dried in air. The l-carboxymethyl- 2,4-dinitropyrrole thus isolated,in almost quantitative yield, is hydrated. It melts at 205207 C., afterslow loss of water of hydration on the melting point stage.

4, Calcd. for C I-I O N C, 33.50; H, 2.34. Found: C, 33.63; H, 2.60.

Example VII.1-(4-methylpiperazinocarbonylmethyl )-2,4-dinitr0pyrr0le Amixture of 10 grams of l-carboxymethyl-Z,4-dinitropyrrole (Example VI)and 75 milliliters of thionyl chloride is boiled under reflux for twohours and then concentrated under vacuum at 60 C. The dark oily residueis dissolved in 50 milliliters of toluene and reconcentrated undervacuum at 60 C. The residue is dissolved in milliliters of methylenechloride and this solution is stirred at 0 while a solution of 5 gramsof N-methyl-piperazine in 40 milliliters of methylene chloride is addedover ten minutes. After being allowed to stand at 25 for fifteen hours,the reaction mixture is shaken with 20 milliliters of 10% aqueous sodiumcarbonate and the methylene chloride layer is separated and dried wtihmagnesium sulfate and then concentrated to a solid residue. This isrecrystallized from ethyl acetate to afford 8.4 grams (61%) of 1(4-methylpiperazino-carbonylmethyl)-2,4- dinitropyrrole as buff flakeswhich melt at -121".

Example VIII .1 (N -pyrr0lidin0carb0nylmethyl 2,4-dinitropyrr0le Amixture of 10 grams of the sodium salt of 2,4-dinitropyrrole, 9.4 gramsof N-chloroacetyl-pyrrolidine, and 40 milliliters of dimethylformamideis boiled -under reflux for fifteen minutes and then poured into 600milli-.

liters of cold 5% aqueous potassium carbonate. The insoluble solid isfiltered off, dried in air and decolorized with charcoal in an acetonesolution. The clear filtrate is evaporated to dryness and the solidresidue is recrystallized from ethyl acetate to alford 12 grams (80%) of1-(N-pyrrolidinocarbonylmethyl) 2,4 dinitropyrrole as pale yellow prismswhich melts at 158-159.

Example IX .-1 ,B-bromoethyl -2-methyl- 5 -n z troim idazo le Fiftymilliliters of thionyl bromide is stirred vigorously and maintained at5060 while a total of 47.5 g. ofl-(B-hydroxyethyl)-2-methyl-5-nitroimidazole is added over a period offifteen minutes. The reaction mixture is then maintained at 80-85 forfifteen minutes, cooled slightly, and poured onto ice, the last of thecontents being rinsed from the reaction vessel with methylene chlorideand a little water. Three hundred milliliters of methylene chloride isadded to the hydrolysis mixture which is then stirred and maintained at5 while enough solid potassium carbonate is added to attain a permanentslightly alkaline pH. The mixture is now filtered and the layers of thefiltrate are separated. The methylene chloride portion is dried withmagnesium sulfate and concentrated under vacuum to a solid yellowresidue which weighs 62.8 grams (96%). This crude [i-bromoethyl compoundis suitable for condensation with the potassium salt of2,4-dinitropyrrole.

Pale yellow granules which melt at -8182 may be obtained byrecrystallization from ether.

Example X.-1-[(2,4-dinitr0pyrryl-I)-etlzyl]-2- methyl-S-nitroimidazole Amixture of 17 grams of the potassium salt of 2,4- dinitropyrrole, 20grams of l-(fi-bromoethyl)-2-methyl- 5-nitroimidazole, and 70milliliters of dimethylformamide is stirred and boiled under reflux forfifteen minutes and then poured into one liter of cold 5% aqueouspotassium carbonate. The insoluble solid is filtered off, dried in airand decolorized with charcoal in an acetone solution.

The pale orange acetone filtrate (about 600 milliliters in volume) isboiled down to a volume of 150 milliliters and chilled at to efifectcrystallization of 16.4 grams of 1- [(2,4 dinitropyrryl 1) ethyl] 2methyl 5 nitroimidazole yellow granules which melt at 186-187.

A second crop of 2.1 grams is obtained on further concentration andchilling of the mother liquor, bringing the total to 18.5 grams, a yieldof The experimental details and physical constants of R ReactionConditions Yield, other compounds of the present invention prepared ac-Dimethylmrmamlde) Percent cording to the methods illustrated in ExamplesI through X above, appear in Tables I, II and III. The constants 1 at 45of compounds not listed in Table III may be found in -04He 83 detailedExamples I through X.

TABLE II.-1-ALKYL-2,4 DINITROPYRROLES, FROM ALKYL HALIDES v KN NOZ RAlkylating Agent Reaction Conditions Yield,

(in Dimethylformamide) Percent I1-C3H7 Il-C3H7Bl 3% hrs. at 135 iso-C Hnis0-C H Br 1 hr. at 145 sec-04119 SeO-C4H9Bl 1% hrs. at 140 11-0513 n-CH11Br hr. at

iso-C5H1 iso-C5H11Br hr. at 140 sec-0 1111 seeCsHuBr hr. at 140 n-onsnan-CuH Br 2 hrs. at GH CHzCHzBr BrCHzCHzCH-gBr 20 min. at 150 01120 0 CH3CHQC 0 CHzCl 5 min. at

CHQCHOHCHZOH CHzOHOHOHCHzCl 50min. at 39 CHQCHQCHBCHZBI'BrCHzCHzCHzCHzBr 10 min. at 145 as CHzCHCHzO O 0 CH3 (Note 1) (Note 2)94 OHzCHa c zo a 0112C ON 0 0101520 ON 0 15 min. at 75 CHZ'C'HB CHQC 0NN-CHa (Note 3) (Note 4) 61 CHzOHz CHQOH,

OHQC ON N C 0 CH3 (Note 5) (Note 6) 25 CH2CHg CH3 CH3 E l l CH2CH2-N/\NBr cnzoH -N N 15 at 70 N02 NOT:

TABLE I.1-ALKYL-2,4DINITROPYRROLES, FROM DIALKYL SULFATES CH2CH2 HN N-CH CHzCHa NOTE 4.--20 hours at 25 in methylene chloride. N OTE 5.Byreaction of R=OH2C 0 01 with NOTE 6.% hour at 5 in methylene chlorid'CHZCHQ HN N-C 00 H TABLE III.-PHYSIGAL CONSTANTS 01]?Afi-qABQlfiYL-ZA-DINITROFYRROLES OF TABLES O gN N Noa Analytical R B.P.0.1mm. M.P.,

degrees Oalcd. Found n-O H1 125-130]. 2 56-57 42. 21C; 4. 5511 42. 26C;4. 65H n-C H9 130-135/. 2 45. 07C; 5. 20H 45. 02C; 5. 3911 150-0411;;137139/. 4 45. 07C; 5. 2011 45. 31C; 5. 18H

sec-C H 125-130]. 1 59-60 45. 07C; 5. 20H 45. 49C; 5. 27H

n-CsHu 146-148]. 18. 49N 18. 62N

150-0 1-1 135-140]. 1 47. 47C; 5. 7711 47. 65C; 5. 72H

seeCsHn 110-115/. 005 47. 57 C; 5. 7711 46. 87C; 5. 7511 n-CaHra125-130]. 002 49. 78C; 6. 2711 49. 71C; 6. H CHgCHgGHzBr 165-170]. 0167-68 30. 43C; 2. 90H 30. 23C; 3. H

CHzCHzCHzCHzB! 180-185]. 003 67-69 32.880; 3. 4511 33. 28C; 3. 6211CHZCO H3 151-152 39. 44C; 3. 31H 39. 88C; 3. 3511 CH2OHOHCHZOH 200-210].02 111-112 36. 37C; 3. 9211 36. 16C; 3. 93H

0 C 0 CH3 CHzCHCHzO COCHa 109-110 41. 91C; 4. 16151 42. 16C; 4. 0011CH2CH5 C1120 ON 0 145-146 42. C; 4. 26H 41. 91C; 4. 15H

o zC z CHzC N r N OHa 120-121 44. 44C; 5. 0911 44. 48C; 5. 1611 CHgCfizCHzCHg OHZC ON NCOCH 215-216 44. 31C; 4. 6511 44. 00C; 4. 68H

CH2 C I 180-187 38. 710; 3.2511 39. 09C; 3. 19H

N Oz

The in vitro trichomonadicidal activity of the compounds of the presentinvention may be demonstrated by a series of tests which establishes theminimal inhibitory concentration of these compounds. Minimal inhibitoryconcentration, as used above, is defined as the minimal concentration ofa trichornonadicidal compound capable of preventing the growth of andkilling T richomonas foetus organisms introduced into a culture medium,capable alone of supporting a vigorous growth of the organisms andcontaining the trichomonadicidal compound to be tested. The culturemedium used in the tests is described in a publication of Kupferberg,Johnson and Sprince, Proceedings of the Society for Experimental Biologyand Medicine, volume 67, pages 3044308, 1948.

In making the tests to determine minimal inhibitory concentrations, 0.05milliliter of a 48-hour culture of Trichomonas foetus is placed in aseries of tubes containing 10 milliliters of the culture medium andincreasing amounts of the compound to be tested. The inoculated culturemedium is then incubated at 37 C. for nine days and examined undermagnification after two, five, seven, and nine days. The minimalinhibitory concentration of the compound tested is that concentration inthe tube in which no viable organisms are present at the ninth dayexamination. If there are no viable organisms present 55 on examinationat a time less than nine days, the concen tration of compound is greaterthan minimal and if there are viable organisms present at the ninth dayexamination, the concentration is less than minimal. The results ofthese tests appear in column 1 of Table IV, wherein 0 the concentrationof the 1-substituted-2,4-dinitropyrroles is expressed in parts permillion.

The toxicity of the compounds of the present invention of T richomonasfoetus is indicated in column 3, under the heading PD In this column,N.P. indicates no protection. Column 4 of Table IV reports thetherapeutic index of these compounds. A therapeutic index of at least 10is preferred by pharmacologists as providing a sufficient margin ofsafety between the LD and the PD TABLE IV.-TRICHOMONADICIDAL ACTIVITY OFl-SUBSTITUTED-2,4-

DINITROPYRROLES 1.1350 P1350 T.I.

n-05H7 100 000 s s0. 0

sec-C1110 10. o 1, 200-1, 250 45 33. 0

Il-CiHll 20. 0 1, 500 50 30 Il-CGHIC! 10. 0 2, 150 140-150 14. 3omcmomBr 10. 0 1, 400 120 12. 0

cmcmomomnr 0. 4 1, 0002, 000 75-100 -20 cmo 0 CH3 10. 0 1, 000 37. 5 27.0

omcnonomon 2. 0 1, 050 -20 52. 0

o o 0 CH5 onion:

CH1 0 ON 1. 0 1, 500 5-7 200 CHzoHt CHrCHz 0 H2O ON N-CHa 10.0 1, 05010.5 100 GHQ-CH2 CH2CH2 omoo-u N-COCHs 10.0 1, 050 07-135 10. 5

CH2CHPN/\] (1) 3, 000 10. 5 300 1 Inactive.

In employing the trichomonad-icides of the present invention for thetreatment of Trichomonas foetus, one 01' more of the active agents areuniformly distributed in a suitable chemotherapeutic vehicle that ischemically compatible with the particular trichomonadicide selected,noninhi'biting with respect to the action of the effective agent upon Trichomonas foetus and essentially non-injurious to the vaginal mucosaunder the conditions of use. The vehicle is preferably of a liquid orsemi-liquid type. Furthermore, since the final preparation should bereadily miscible with vaginal fluids, the vehicles, whether hydrous oranhydrous, are preferably water-miscible or water-dispersi'ble. Thecompositions of this invention may 'be in the form of suppositories, ifdesired.

The foregoing criteria for a vehicle in which the compounds of thepresent invention are incorporated may be met by a large number ofsemi-liquid chemotherapeutic vehicles that are well known in the art.Thus, for example, the vehicle may comprise semi-liquids that arecolloidal in nature, especially those that are viscous and/ ormucilaginous in character. Such vehicles are particularly suitable foruse in topical treatment of Trichomonas foetus because of their inherentgelatinous and miscible nature which affords prolonged contact betweenthe 1- substituted-Z,4-dinitropyrrole and the infecting organism.

In order to disclose more clearly the manner of forumlating thecompounds of the present invention to topical application, severalspecific examples will hereinafter be described in considerable detail.

Example XI Deionized water 75.80

The trichomonadicidal formulations of Examples XI through XIII areprepared according to the following general procedure in which twoinitial solutions are mixed to make the formulation, all the parts beinggiven by weight. To prepare Solution A, dissolve the parahydroxy-benzoicacid methyl ester in about two-thirds of the hot deionized water, coolto about 170 F., and, while stirring, add the gel-forming ingredient andglycerine or propylene glycol. To prepare Solution B, add thetriohomonadicidal agent to the remainder of the deionized water, andadjust the pH to the desired. value. The formulation is prepared byadding Solution B to Solution A in a slow stream with good stirring;stirring is continued for at least one hour.

Certain compounds of the present invention have also been found to beeffective against enterohepatitis (blackhead) when administered byadmixture, suspension, or dispersion in the food and/ or drink normallypartaken by turkeys, such as grain, mash, scratch, water or otherliquids.

The general range of concentration of thel-substituted-2,4-dinitropyrrole in the total substance is from about0.05% or less to about 1%. The optimal concentration for eifectivetherapy is in the range from about 0.05% to about 0.2% of the total foodor drinking water. With these optimal concentrations, the daily drugintake for infected birds varies from about 20 milligrams of drug perkilogram of body weight to about 400 milligrams of drug per kilogram ofbody 12 weight. In general, the precise dosage depends on the particularcompound and the severity of the infection. Many of the compounds of thepresent invention may be administered in the concentrations indicatedabove with little or no toxic effect.

Various changes and modifications of the invention may be made and tothe extent that such variations incorporate the spirit of thisinvention, they are intended to be included within the scope of theappended claim.

What is claimed is:

A compound of the formula I-C-NO:

CHa-C I NCH No references cited.

